Identification of the modulatory Ca 2+ -binding sites of acid-sensing ion channel 1a.

Acid-sensing ion channels (ASICs) are neuronal Na ⁺ -permeable ion channels activated by extracellular acidification. ASICs are involved in learning, fear sensing, pain sensation and neurodegeneration. Increasing the extracellular Ca ²⁺ concentration decreases the H ⁺ sensitivity of ASIC1a, suggesting a competition for binding sites between H ⁺ and Ca ²⁺ ions. Here, we predicted candidate residues for Ca ²⁺ binding on ASIC1a, based on available structural information and our molecular dynamics simulations. With functional measurements, we identified several residues in cavities previously associated with pH-dependent gating, whose mutation reduced the modulation by extracellular Ca ²⁺ of the ASIC1a pH dependence of activation and desensitization. This occurred likely owing to a disruption of Ca ²⁺ binding. Our results link one of the two predicted Ca ²⁺ -binding sites in each ASIC1a acidic pocket to the modulation of channel activation. Mg ²⁺ regulates ASICs in a similar way as does Ca ²⁺ . We show that Mg ²⁺ shares some of the binding sites with Ca ²⁺ . Finally, we provide evidence that some of the ASIC1a Ca ²⁺ -binding sites are functionally conserved in the splice variant ASIC1b. Our identification of divalent cation-binding sites in ASIC1a shows how Ca ²⁺ affects ASIC1a gating, elucidating a regulatory mechanism present in many ion channels.